Spontaneous hydrolytic deamination of DNA bases represents a considerable mutagenic threat to all organisms, particularly those living in extreme habitats. Cytosine is readily deaminated to uracil, which base pairs with adenine during replication, and most organisms encode at least one uracil DNA glycosylase (UDG) that removes this aberrant base from DNA with high efficiency. Adenine deaminates...

Base moieties in DNA are spontaneously threatened by naturally occurring chemical reactions such as deamination, hydrolysis and oxidation. These DNA modifications have been considered to be major causes of cell death, mutations and cancer induction in organisms. Organisms have developed the DNA base excision repair pathway as a defense mechanism to protect them from these threats. DNA glycosyla...

A revival of interest has taken place in recent years in the possibility of interference with mam malian nucleic acid biosynthesis by means of an tagonists of the free pyrimidines. Early studies appeared to show that, while orotic acid (1, 48) and the sugar derivatives of the pyrimidines (19, 20) are extensively incorporated into both RNA and DNA, the incorporations of free uracil, cytosine, an...

Schizosaccharomyces pombe Δura4 cells lyse when grown on YPD medium. A S. pombe non-essential gene deletion library was screened to determine suppressors of the lysis phenotype. Deletion of the pub1 gene, which encoded E3 ubiquitin ligase, strongly suppressed cell lysis in Δura4 cells. The Δpub1 cells displayed high sensitivity to 5-fluorouracil, a toxic analog of uracil, and this sensitivity w...

The Hunter (1) procedure was originally developed for the calorimetric d&&ion of minute amounts of thymine. In this procedure thymine is coupled under air with p-diazobenzenesulfonic acid in carbonate solut.ion, and a red color is subsequently produced by the addition of sodium hydroxide and hydroxylamine. Thymine was determined by this means in hydrolysates of dcsoxypentose nucleic acids (DNA)...

Gene-targeted mice deficient in the evolutionarily conserved uracil-DNA glycosylase encoded by the UNG gene surprisingly lack the mutator phenotype characteristic of bacterial and yeast ung(-) mutants. A complementary uracil-DNA glycosylase activity detected in ung(-/-) murine cells and tissues may be responsible for the repair of deaminated cytosine residues in vivo. Here, specific neutralizin...

The base-pairing interactions of promutagenic O6-methylguanine (O6-MeGua) with cytosine and thymine in deuterated chloroform were investigated by 1H NMR spectroscopy. Nucleosides were derivatized at hydroxyl positions with triisopropylsilyl groups to obtain solubility in nonaqueous solvents and to prevent the ribose hydroxyls from forming hydrogen bonds. We were able to observe hydrogen-bonding...

Rat liver nuclei were separated into two fractions, the nucleolus and the nucleoplasm, by a combined salt-enzymic extraction. RNA was purified from both sources and analysed on sucrose density gradients. In both fractions a prominent heterogeneous RNA class with mean sedimentation coefficient of 18S was found. This material was analysed by measuring the rate of reaction with formaldehyde, the u...

DNA glycosylases UNG and SMUG1 excise uracil from DNA and belong to the same protein superfamily. Vertebrates contain both SMUG1 and UNG, but their distinct roles in base excision repair (BER) of deaminated cytosine (U:G) are still not fully defined. Here we have examined the ability of human SMUG1 and UNG2 (nuclear UNG) to initiate and coordinate repair of U:G mismatches. When expressed in Esc...

Expression of the bacterial gene for cytosine deaminase (CD; EC 3.5.4.1) in mammalian cells was evaluated as a negative selection system or suicide vector for potential use in gene transfer studies and therapies. Mammalian cells, unlike certain bacteria and fungi, do not contain the enzyme CD and do not ordinarily metabolize cytosine to uracil. Nor do they metabolize the innocuous compound 5-fl...